This study aimed to determine the prevalence of vesicular stomatitis virus (VSV) in buffaloes in the state of Paraíba, Northeastern Brazil. The study was carried out in 14 herds in the municipalities of Alagoa Nova, Areia, Campina Grande, Guarabira, Juripiranga, Santa Helena, Sapê, Rio Tinto, Santana dos Garrotes, Itatuba, Solânea, and Cacimbas. The studied population was formed by buffalo females reared for meat and milk, comprising of mixed and Murrah breeds, at least 24 months of age. For the serological diagnosis of VSIV-3, the virus-neutralization test (VN) was performed, using the VSIV-3 isolate 2013 São Bento/Paraíba. Real prevalence was 2.6% for VSIV-3, with antibody titers ranging from 160 to 1280. This is the first study to characterize VSV circulation in the buffalo population in Northeastern Brazil, where infection is considered endemic; some aspects of virus maintenance are not fully understood, such as the role of reservoirs in endemic areas. The identification of seroreactive animals in this study demonstrates the circulation of VSIV-3 in the buffalo species. Reports of virus isolation in this species have not yet been described, which suggests the need for investigating the role of buffaloes in vesicular stomatitis epidemiology.